Mem Inst Oswaldo Cruz, Rio de Janeiro, Vol. 93(3): 391-398, May/Jun. 1998
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HIV Specific Humoral Immune Response in Rio de Janeiro, Brazil V Bongertz/+, CI Costa, ML Guimarães, MFG Soares-da-Costa, B Grinsztejn*, The HEC/Fiocruz AIDS Clinical Research Group*/++, FI Bastos**, JH Pilotto***, EC João Filho****, R Loureiro*****, P Chequer******, PR Telles*******, B Galvão-Castro********, MG Morgado Laboratório de AIDS e Imunologia Molecular, Departamento de Imunologia, *Hospital Evandro Chagas, Instituto Oswaldo Cruz, Av. Brasil 4365, 21045-900 Rio de Janeiro, RJ, Brasil **Centro de Informações em Ciência e Tecnologia, Fiocruz, RJ ***Hospital Geral de Nova Iguaçu, RJ **** Hospital dos Servidores do Estado, RJ *****Secretaria da Saúde e Meio Ambiente, Porto Alegre, RS ******Programa Nacional DST/ AIDS, MS, Brasília, DF *******NEPAD, Universidade Estadual do Rio de Janeiro ********Laboratório Avançado de Saúde Pública, Centro de Pesquisas Gonçalo Moniz, Fiocruz, Salvador, BA, Brasil
Efforts to characterize HIV-1 polymorphism and anti-HIV immune response are being made in areas where anti-HIV/AIDS vaccines are to be employed. Anti-HIV-1 humoral immune response is being studied in infected individuals resident in Rio de Janeiro, in distinct cohorts involving recent seroconvertors, pregnant women or intravenous drug users (IDU). Comparative analyses of specificity of antibody response towards epitopes important for anti-HIV-1 immune response indicate quantitative differences between cohorts, with an exceptionally strong response in IDUs and weakest response in pregnant women. However, a comparative analysis between pregnant women cohorts from Rio de Janeiro and Rio Grande do Sul indicated an even lower response (with exception of the anti-V3-C clade peptide recognition) for the southern cohort. Studies analysing the immune function of the humoral response indicate a quite elevated occurrence of antibodies capable of neutralizing heterologous primary HIV-1 isolates from Rio de Janeiro. Attempts to correlate seroreactivity with HIV-1 neutralization with respect to HIV-1 polymorphism were not very successfull: while the Brazilian B clade B” variant could be recognized by binding assays, no significant distinction of HIV-1 clades/variants was observed in viral neutralization assays. Key words: HIV-1 - antibody specificity - neutralization - genotypes
Attempts to develop anti-HIV/AIDS vaccines have been checked by the great genetic variability of HIV-1 (Bloom 1996). Genotyping studies have subdivided HIV-1 into several subtypes, while serotyping analyses have not lead to a definition of distinct HIV-1 serotypes (Moore & Trkola 1997). A clustering of “serospecificities” based on binding of antibodies to synthetic peptides corresponding to genotypic subtypes indicate a partial correspondence between genotype and serotype
This work was supported by PIAF/Fiocruz, MS, CNPq 529022/95-6 and UNDP/World Bank 063/94 +Corresponding author. Fax: +55-21-280.1486. E-mail:
[email protected] ++The HEC/Fiocruz AIDS Clinical Research Group: VGV Santos, VC Rolla, S Cavalcante, MCG Galhardo, MRC Guimarães, R Khalili, R Valles de Souza Received 11 March 1998 Accepted 16 April 1998
(Moore et al. 1996, Simon et al. 1996). However, analyses of the functional HIV neutralizing humoral response has not indicated a correlation with HIV-1 genotypes (Weber et al. 1996) but a “grouping” of sera with similar neutralization potency and cross-clade activity (Trkola et al. 1995, Kostrikis et al. 1996, Nyambi et al. 1996). The need for an anti-AIDS vaccine concomitant to HIV-1 variability motivates studies of viral polymorphism in the different geographic regions with high prevalence of HIV infection, in parallel to studies on specific human immune response to the various clades in the different areas. Identification of specific immune reactivities (Fenyö et al. 1996) and analyses of its potential protectivity against development of immune deficiency in HIV infection is needed for development and evaluation of effective vaccine candidates. This report presents results of several studies of humoral immune response against HIV-1 in Rio de Janeiro, Brazil, in different patient cohorts to allow comparative evaluations.
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Anti-HIV Antibodies in Brazil V Bongertz et al.
MATERIALS AND METHODS
Patients - Data is presented from different patient groups: (a) Vertical HIV transmission group (TV-RJ), accompanied at the Hospital Evandro Chagas, Instituto Oswaldo Cruz, RJ; the Hospital Geral de Nova Iguaçu, RJ and the Hospital dos Servidores do Estado, RJ, comprised of 125 HIV1 infected pregnant women and women that had given birth recently; (b) 30 individuals from an injecting drug user (IDU) cross-sectional study, comprised of a mostly male group (97%), of the “Projeto Brasil”, Centro de Informações em Ciência e Tecnologia, Fiocruz, RJ, and NEPAD, Universidade Estadual do Rio de Janeiro; (c) 85 asymptomatic HIV-1 infected individuals resident in Rio de Janeiro (RN-RJ), Hospital Evandro Chagas; (d) a cohort of 51 pregnant asymptomatic HIV-1 infected women resident in Rio Grande do Sul (TV-RS), accompanied by the Secretaria da Saúde e Meio Ambiente, Porto Alegre, RS. Seroreactivity - Duplicates of sequentially diluted heat inactivated plasma were incubated with different biotinylated synthetic peptides bound to multiwell plates precoated with streptavidin. Specific reactivity was assessed (including 8M urea washes) by peroxidase-conjugated anti-human-IgG binding and H2O2/TMB revelation (Bastos et al. 1997). Reactivity was assessed as dilution titer reaching at least twice the mean binding of the two control plasma included in each assay plate. The variation of total numbers of plasma analyzed derives from exclusion of data from doubtful analyses. Neutralization assay - Supernatants from primary co-cultures collected between the 7th and 21st day of co-culture were used as viral stocks. The standard assay employing pre-incubation of viral dilutions with heat inactivated plasma followed by addition of phytohemagglutinin-activated normal human PBMC (Albert et al. 1990, Bongertz et al. 1997). Changes of at least 90% of the medium after 24 and 48 hr were carried out in order to eliminate eventually present anti-p24 antibodies in the plasma used (Albert et al. 1993). Evaluation of neutralization was carried out by measurement of the HIV-1 p24 antigen (DuPont HIV-1 antigen assay) in the culture supernatants on day 7. Neutralization was considered positive when a reduction of at least 75% of viral input was detected as measured by p24 concentration (Bongertz et al. 1997). Heteroduplex mobility assay (HMA) - The test was carried out according to Delwart et al. (1993) using the reagents indicated. Differentiation between B subtype and the B” variant of subtype B was carried out using analysis of Fok 1 restriction enzyme fragments by agarose gel electrophoresis (Morgado et al. 1996).
Statistical analyses - Fisher’s exact test was used for statistical evaluation (significance defined as p
